title: Hypoxia alters phosphorylation status of insulin-like growth factor (IGF)-binding protein-1 and attenuates biological activities of IGF-I in HepG2 cell cultures.
abstract: OBJECTIVE
  Insulin-like growth factor (IGF)-I is known to stimulate fetal growth. One of the IGF-binding proteins, IGFBP-1, suppresses IGF-I activity, and thereby inhibits fetal growth. Because hypoxic stress in the uterus is known to cause fetal growth restriction, we examined the effects of hypoxia on IGFBP-1 production and phosphorylation status.
  
  
  METHODS
  Because liver is a main IGFBP-1 production site in the fetus, we used a hepatoma cell line, HepG2 cells, that secrete a large amount of IGFBP-1, express IGF-I receptors and model fetal liver metabolism in vitro. IGFBP-1 was analyzed by sodium dodecylsulfate polyacrylamide gel electrophoresis (PAGE) following immunoblotting, and IGFBP-1 phosphorylation status was analyzed by native PAGE following immunoblotting.
  
  
  RESULTS
  Total concentrations of IGFBP-1 in media were higher and the highly phosphorylated isoforms were dominant in low oxygen conditions. Phosphorylation of IGF-I receptor by IGF-I was attenuated in low oxygen conditions. IGF-I-induced phosphorylation of insulin receptor substrate-1 (IRS-1) was attenuated in low oxygen conditions as well. However, attenuated phosphorylation of IGF-I receptor and IRS-1 were not observed in low oxygen conditions if the cells were stimulated with LR³IGF-I that has a similar binding affinity to IGF-I receptor but much less binding affinity to IGFBP-1 compared to those of native IGF-I. While IGF-I-induced cell proliferation was also inhibited in low oxygen conditions, LR³IGF-I-stimulated cell proliferation was not inhibited. These findings indicate that low oxygen conditions inhibit IGF-I action by increasing IGFBP-1, especially phosphorylated IGFBP-1, which inhibits IGF-I action.
  
  
  CONCLUSIONS
  This study has indicated that hypoxia-induced IGFBP-1 production in the fetus may be a conserved physiological mechanism for restricting IGF-I-stimulated fetal growth.
authors.affiliation: Department of Obstetrics and Gynecology, School of Medicine, Kyorin University, Tokyo, Japan.
authors.forename: Tomonori
authors.initials: T
authors.lastname: Tsunawaki
authors.forename: Keiji
authors.initials: K
authors.lastname: Sakai
authors.forename: Mai
authors.initials: M
authors.lastname: Momomura
authors.forename: Yuichi
authors.initials: Y
authors.lastname: Wachi
authors.forename: Yukiko
authors.initials: Y
authors.lastname: Matsuzawa
authors.forename: Mitsutoshi
authors.initials: M
authors.lastname: Iwashita
mesh_terms: D015687:Cell Hypoxia; D056945:Hep G2 Cells; D022781:Hepatocytes / Q000378:metabolism*; D006801:Humans; D055504:Insulin Receptor Substrate Proteins / Q000378:metabolism; D018970:Insulin-Like Growth Factor Binding Protein 1 / Q000378:metabolism*; D007334:Insulin-Like Growth Factor I / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D017526:Receptor, IGF Type 1 / Q000378:metabolism; D015398:Signal Transduction; D015854:Up-Regulation
chemical_list: C583557:IGF1 protein, human; C032231:IGFBP1 protein, human; C526292:IRS1 protein, human; D055504:Insulin Receptor Substrate Proteins; D018970:Insulin-Like Growth Factor Binding Protein 1; D007334:Insulin-Like Growth Factor I; D017526:Receptor, IGF Type 1
keywords: fetal growth; hypoxia; insulin-like growth factor-binding protein-1